The main techniques and approaches of biochemical cytology, resting largely on the combined use of cell fractionation, biochemical analysis, and light and electron microscopy, are applied to a detailed analysis of the activities and properties of lymphocytes in an in vitro cytotoxicity system. We will study the biochemical and cytological events accompanying the differentiation of uncommitted rat lymphocytes into specific cytotoxic cells, and we will follow the kinetics of this process by autoradiography. Special attention will be given to the properties of the large cytotoxic lymphocytes, purified by means of zonal centrifugation. The possibility that these cells use lysosomal enzymes for killing target cells will be examined, and detailed studies will be performed to characterize the surface proteins on the plasma membrane by the lactoperoxidase method and to compare the results with those obtained on small T-lymphocytes.